1. Technical Field
The present invention relates to methods of extraction of oil from biomass, and in particular, algae. The present invention further relates to methods of extraction from algae biomass containing a controlled level of moisture.
2. Background Art
The mechanism of solvent extraction from oil-bearing biomass has been well studied, and engineers have applied leaching theory, diffusion theory, soaking theory, and viscous capillary flow to explain extraction kinetics and design efficient extractors (L. A. Johnson, 1997. “Theoretical, Comparative, and Historical Analyses of Alternative Technologies for Oilseeds Extraction”, p. 4-47. In P. J. Wan and P. J. Wakelyn (ed.), Technology and solvents for extracting oilseeds and nonpetroleum oil. The American Oil Chemists Society, Champaign, Ill.). In commercial operations, most oilseeds undergo significant pre-extraction processing to improve extraction efficacy. For example, seeds are generally heated, cracked, and flaked, thereby rupturing cell walls and making oil more available to extraction solvents. Furthermore, flaking reduces the distance over which the oil must be transferred to dissolve in the solvent. Since the transfer mechanism is highly dependent on capillary flow, and to some extent on the viscosity of the solvent and miscella (oil-solvent mixture), feedstock preparation and solvent choice are large factors in extraction yields. In fact, flake thickness is often regarded as the most important factor in extraction efficiency, suggesting that feedstock preparation is a crucial part of the process that cannot be neglected when considering extracting lipids from microalgae (P. J. Wan and P. J. Wakelyn (ed.), Technology and solvents for extracting oilseeds and nonpetroleum oil. The American Oil Chemists Society, Champaign, Ill., 1997).
In reference to soybeans, Johnson (1997) notes that oil in unruptured cells diffuses out of the cell by osmosis, which is a very slow process and whose rate depends on the molecular size of the oil and solvent. Analogous considerations should be made for algae biomass: intact algae cells with durable cell walls represent a barrier to extraction of intracellular compounds, such as triglycerides, fatty acids and other lipids within the intact cell. Cell disruption by homogenization, sonication, high pressure, and pure solvents have all been investigated and reported to significantly influence the amount of recovered oil. Lipid solubility is a crucial factor in extraction procedures because success is predicated on finding a solvent system that will dissolve the lipids of interest while overcoming the interactions between lipids and their surroundings (S. J. Iverson et al., “Comparison of the Bligh and Dyer and Folch methods for total lipid determination in a broad range of marine tissue”, Lipids 36 (11), (2001) 1283-1287). Generally, the solubility of pure lipids depends on their polarity and that of the solvent. Triglycerides are very soluble in non-polar solvents such as hexane, cyclohexane, and toluene, as well as slightly more polar solvents like chloroform (W. W. Christie (2003) Lipid analysis: isolation, separation, identification and structural analysis of lipids”). The solubility of triglycerides in polar solvents, such as alcohols, is very low. As alcohol chain-length increases and fatty acid chain length decreases, triglycerides become more soluble in alcohols; however, the surprising efficacy of ethanol in extracting algal lipids in the C16 to C22 chain length is clearly not obvious to someone ordinarily skilled in the art. In contrast to simple lipids, polar complex lipids have low solubility in hydrocarbon solvents, but can dissolve readily in chloroform, methanol, and ethanol (W. W. Christie (2003) Lipid analysis: isolation, separation, identification and structural analysis of lipids”). These observations teach that different solvents are needed to efficiently extract the different lipids within the algal biomass.
Recognizing the need to balance non-polar solvents capable of dissolving simple (neutral) lipids with polar solvents capable of extracting complex lipids, scientists and engineers have developed procedures using mixtures of solvents such as 2:1 blends of chloroform and methanol to quantitatively recover almost all major lipid classes from a variety of samples. These procedures, originally developed by Bligh and Dyer, Folch and others underlie all of the current knowledge regarding lipid content (and lipid extraction) in microalgae. These technologies are then applied to biomass that is pretreated (e.g. homogenization in a blender) as described above. These methods are unsuitable for commercial application due to the impracticality of using and recycling solvent mixtures, the toxicity of the preferred solvents (such as chloroform, hexane, and methanol) and their unacceptability in key applications such as nutraceuticals, pharmaceuticals and nutrition. These methods are unsuitable for extracting valuable oils such as omega-3 oils from phospholipids and glycolipids. These methods also use mechanical means requiring excessive energy (resulting in unfavorable energy balance and high costs plus degradation of the resulting algae fractions) or high temperatures that decrease the value of the products due to oxidation, isomerization, hydrolysis, degradation, or other pathways to decomposition.
There are several processes currently used that involve dry biomass or drying wet biomass to an extent. For example, U.S. Pat. No. 6,441,208 to Biil, et al. discloses a microbial polyunsaturated fatty acid (PUFA)-containing oil with a high triglyceride content (greater than 90%), and hence essentially devoid of polar lipids, and a high oxidative stability. In addition, a method is disclosed of the recovery of such oil from a microbial biomass derived from a pasteurized fermentation broth, wherein the microbial biomass is subjected to extrusion to form granular particles, dried and the oil then extracted from the dried granules using an appropriate solvent. The '208 patent forms granules out of dry biomass powder (the biomass can be algae). Solvent is then contacted with the granules, and the solvent can be ethanol or other alcohols to extract compounds/oils.
U.S. Pat. No. 7,868,195 to Fleischer, et al. discloses centrifuging a wet algal biomass to increase a solid content of the wet algal biomass to between approximately 10% and 40% to result in a centrifuged algal biomass, mixing the centrifuged algal biomass with an amphiphilic solvent to result in a mixture, heating the mixture to result in a dehydrated, defatted algal biomass, separating the amphiphilic solvent from the dehydrated, defatted algal biomass to result in amphiphilic solvent, water and lipids, evaporating the amphiphilic solvent from the water and the lipids, and separating the water from the lipids. The amphiphilic solvent may be selected from a group consisting of acetone, methanol, ethanol, isopropanol, butanone, dimethyl ether, and propionaldehyde. Other exemplary methods include filtering a wet algal biomass through a membrane to increase a solid content of the wet algal biomass to between approximately 10% and 40% to result in a filtered algal biomass. Separation can be performed by membrane filtration or centrifugation.
U.S. Pat. No. 8,153,137 to Kale discloses a method of isolating nutraceuticals products from algae. A method of isolating carotenoids and omega-3 rich oil from algae includes dewatering substantially intact algal cells to make an algal biomass and adding a first ethanol fraction to the algal biomass. The method also includes separating a first substantially solid biomass fraction from a first substantially liquid fraction comprising proteins and combining the first substantially solid biomass fraction with a second ethanol fraction. The method further includes separating a second substantially solid biomass fraction from a second substantially liquid fraction comprising polar lipids and combining the second substantially solid biomass fraction with a third ethanol solvent fraction. The method also includes separating a third substantially solid biomass fraction from a third substantially liquid fraction comprising neutral lipids, wherein the third substantially solid biomass fraction comprises carbohydrates and separating the neutral lipids into carotenoids and omega-3 rich oil. This is an energy intensive process due to the many step-wise separation steps required and is generally not a practical process. This process is a fractionation process to obtain proteins, polar lipids, carbohydrates, carotenoids, chlorophyll, and omega-3 fatty acids.
There remains a need for a method of extracting valuable algae oils from algae without causing damage to their structure, composition or commercial value (or that of the residual extracted algae meal) and without using solvents that are unacceptable in the above markets, since residual levels of said solvents diminish the value of the final products, and in many cases make them unacceptable in that market segment. There further remains a need for a method of extraction that is energy and cost efficient.